Imidazolophthalazine derivatives as ligands for gabaa receptors

ABSTRACT

A class of substituted imidazolo[5,1-a]phthalazine derivatives as ligands for GABA A  receptors which are partial or full inverse agonists of an α5 receptor subunit while being relatively free of activity at α1 and/or α2 and/or α3 receptor subunit binding sites; they are therefore of benefit as a medicament for enhancing cognition but with the reduction or elimination of procon-vulsant activity.

[0001] The present invention relates to a class of substitutedimidazolophthalazine derivatives and to their use in therapy. Moreparticularly, this invention is concerned with substitutedimidazolo[5,1-a]phthalazine derivatives which are ligands for GABA_(A)receptors containing the α5 subunit and are therefore useful in therapywhere cognition enhancement is required.

[0002] Receptors for the major inhibitory neurotransmitter,gamma-aminobutyric acid (GABA), are divided into two main classes: (1)GABA_(A) receptors, which are members of the ligand-gated ion channelsuperfamily; and (2) GABA_(B) receptors, which may be members of theG-protein linked receptor superfamily. Since the first cDNAs encodingindividual GABA_(A) receptor subunits were cloned the number of knownmembers of the mammalian family has grown to thirteen (six α subunits,three β subunits, three γ subunits and one δ subunit). It may be thatfurther subunits remain to be discovered; however, none has beenreported since 1993.

[0003] Although knowledge of the diversity of the GABA_(A) receptor genefamily represents a huge step forward in our understanding of thisligand-gated ion channel, insight into the extent of subtype diversityis still at an early stage. It has been indicated that an α subunit, βsubunit and a γ subunit constitute the minimum requirement for forming afully functional GABA_(A) receptor expressed by transiently transfectingcDNAs into cells. As indicated above, a δ subunit also exists, but isapparently uncommon in the native receptor.

[0004] Studies of receptor size and visualisation by electron microscopyconclude that, like other members of the ligand-gated ion channelfamily, the native GABA_(A) receptor exists in pentameric form. Theselection of at least one α, one β and one γ subunit from a repertoireof thirteen allows for the possible existence of more than 10,000pentameric subunit combinations. Moreover, this calculation overlooksthe additional permutations that would be possible if the arrangement ofsubunits around the ion channel had no constraints (i.e. there could be120 possible variants for a receptor composed of five differentsubunits).

[0005] Receptor subtype assemblies which do exist include α1β2γ2,α2β2/3γ2, α3βγ2/3, α2βγ1, α5β3γ2/3, α6βγ2, α6βδ and α4βδ. Subtypeassemblies containing an α1 subunit are present in most areas of thebrain and account for over 40% of GABA_(A) receptors in the rat. Subtypeassemblies containing α2 and α3 subunits respectively account for about25% and 17% of GABA_(A) receptors in the rat. Subtype assembliescontaining an α5 subunit are primarily hippocampal and represent about4% of receptors in the rat.

[0006] A characteristic property of some GABA_(A) receptors is thepresence of a number of modulatory sites, of which the most explored isthe benzodiazepine (BZ) binding site through which anxiolytic drugs suchas diazepam and temazepam exert their effect. Before the cloning of theGABA_(A) receptor gene family, the benzodiazepine binding site washistorically subdivided into two subtypes, BZ1 and BZ2, on the basis ofradioligand binding studies. The BZ1 subtype has been shown to bepharmacologically equivalent to a GABA_(A) receptor comprising the alsubunit in combination with β2 and γ2. This is the most abundantGABA_(A) receptor subtype, representing almost half of all GABA_(A)receptors in the brain.

[0007] A number of dementing illnesses such as Alzheimer's disease arecharacterised by a progressive deterioration in cognition in thesufferer. It would clearly be desirable to enhance cognition in subjectsdesirous of such treatment, for example for subjects suffering from adementing illness.

[0008] It has been reported by McNamara and Skelton in Psychobiology,21:101-108, that the benzodiazepine receptor inverse agonist β-CCMenhanced spatial learning in the Morris watermaze. However, β-CCM andother conventional benzodiazepine receptor inverse agonists areproconvulsant which makes it clear that they cannot be used as cognitionenhancing agents in humans.

[0009] However, we have now discovered that it is possible to obtainmedicaments which have cognition enhancing effects which may be employedwith less risk of proconvulsant effects previously described withbenzodiazepine receptor partial or fill inverse agonists.

[0010] It has now been discovered that use of an α5 receptor partial orfull inverse agonist which is relatively free of activity at α1 and/orα2 and/or α3 receptor binding sites can be used to provide a medicamentwhich is useful for enhancing cognition but in which proconvulsantactivity is reduced or eliminated. Inverse agonists at α5 which are notfree of activity at α1 and/or α2 and/or α3 but which are functionallyselective for α5 can also be used. Inverse agonists which are bothselective for α5 and are relatively free of activity at α1, α2 and α3receptor binding sites are preferred.

[0011] The present invention provides a compound of formula I, or apharmaceutically acceptable salt thereof:

[0012] wherein

[0013] L is O, S or NR^(n) where R^(n) is H, C₁₋₆alkyl orC₃₋₆cycloalkyl;

[0014] X is a 5-membered heteroaromatic ring containing 1, 2, 3 or 4heteroatoms independently chosen from oxygen, nitrogen and sulphur, atmost one of the heteroatoms being oxygen or sulphur, or a 6-memberedheteroaromatic ring containing 1, 2 or 3 nitrogen atoms, the 5- or6-membered heteroaromatic ring being optionally fused to a benzene orpyridine ring and the heteroaromatic ring being optionally substitutedby R^(x) and/or R^(y) and/or R^(z), where R^(x) is halogen, R³, OR³,OCOR³, NR⁴R⁵, NR⁴COR⁵, OH, tri(C₁₋₆alkyl)silylC₁₋₆alkoxyC₁₋₄alkyl, CN orR⁶, R^(y) is halogen, R³, OR³, OCOR³, NR⁴R⁵, NR⁴COR⁵, NR⁴R⁵(C₁₋₆)alkylor CN and R^(z) is R³, OR³ or OCOR³, providing that when X is a pyridinederivative, the pyridine ring is optionally in the form of the N-oxide,and providing that when X is a tetrazole derivative it is protected by aC₁₋₄alkyl group;

[0015] Y is optionally branched C₁₋₄alkylene optionally substituted byan oxo group or Y is a group (CH₂)_(j)O wherein the oxygen atom isnearest the group X and j is 2, 3 or 4;

[0016] Z represents a phenyl ring, a 5-membered heteroaromatic ringcontaining 1, 2 or 3 heteroatoms independently selected from oxygen,nitrogen and sulphur, at most one of the heteroatoms being oxygen orsulphur, or a 6-membered heteroaromatic ring containing 1, 2 or 3nitrogen atoms with the exception of pyrazine, any of which rings may beoptionally substituted with one or more substituents selected fromhalogen, R³, OR³, OCOR³, NR⁴R⁵, NR⁴R⁵(C₁₋₆)alkyl, NR⁴R⁵CO,NR⁴R⁵CO(C₁₋₆)alkyl, CN, cyano(C₁₋₆)alkyl or R⁶;

[0017] R³ represents C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkyl, C₃₋₆cycloalkyl,C₃₋₆cycloalkyl(C₁₋₆)alkyl, cyano(C₁₋₆)alkyl, hydroxyC₁₋₆alkyl and R³ isoptionally mono, di- or tri-fluorinated;

[0018] R⁴ and R⁵ are each independently hydrogen, C₁₋₆alkyl,C₂₋₆alkenyl, C₂₋₆a ynyl, C₃₋₆cycloalkyl or CF₃ or R⁴ and R⁵, togetherwith a nitrogen atom to which they are commonly attached, form a 4-7membered heteroaliphatic ring containing the said nitrogen atom andoptionally one other heteroatom selected from O, N and S; and

[0019] R⁶ is C₆₋₁₀aryl, C₆₋₁₀aryl (C₁₋₆)alkyl, heteroaryl orheteroaryl(C₁₋₆)alkyl, where heteroaryl is a five-membered aromatic ringcontaining one, two, three or four nitrogen atoms or one oxygen atom orsulphur atom and optionally one or two nitrogen atoms or a six-memberedaromatic ring containing one, two or three nitrogen atoms, and R⁶ isoptionally substituted by one, two or three substituents independentlychosen from halogen atoms and C₁₋₄alkyl, C₂₋₄alkenyl, C₂₋₄alkynyl,C₁₋₄alkoxy, C₂₋₄alkenyloxy and C₂₋₄alkynyloxy groups each of whichgroups is unsubstituted or substituted by one, two or three halogenatoms.

[0020] As used herein, the expression “C₁₋₆alkyl” includes methyl andethyl groups, and straight-chained and branched propyl, butyl, pentyland hexyl groups. Particular alkyl groups are methyl, ethyl, n-propyl,isopropyl and t-butyl. Derived expressions such as “C₁₋₄alkyl”,“C₂₋₄alkenyl”, “C₂₋₆alkenyl”, “hydroxyC₁₋₆alkyl”, “C₂₋₄alkyl” and“C₂₋₆alkynyl” are to be construed in an analogous manner.

[0021] The expression “C₃₋₆cycloalkyl” as used herein includes cyclicpropyl, butyl, pentyl and hexyl groups such as cyclopropyl andcyclohexyl.

[0022] The expression “C₃₋₆ cycloalkyl(C₁₋₆)alkyl” as used hereinincludes cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl andcyclohexylmethyl.

[0023] Typical C₄₋₇ cycloalkenyl groups include cyclobutenyl,cyclopentenyl and cyclohexenyl.

[0024] The expression “C₁₋₄alkylene” as used herein refers to alkanediylgroups of up to 4 carbon atoms in which the unsatisfied valencies resideon the same carbon atom or on different carbon atoms.

[0025] Typical C₆₋₈ bicycloalkyl groups include bicyclo[2.1.1]hexyl andbicyclo[2.2.1]heptyl.

[0026] Typical aryl groups include phenyl and naphthyl, preferablyphenyl.

[0027] The expression “aryl(C₁₋₆)alkyl” as used herein includes benzyl,phenylethyl, phenylpropyl and naphthylmethyl.

[0028] Suitable heterocycloalkyl groups include azetidinyl,pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl and thiomorpholinylgroups.

[0029] Unless otherwise specified, 5- and 6-membered heteroaromaticrings shall include pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl,pyrrolyl, furyl, thienyl, pyrazolyl, oxazolyl, isoxazolyl, thiazolyl,isothiazolyl, imidazolyl, oxadiazolyl, triazolyl and thiadiazolyl groupsand benzo-fused analogues thereof A suitable 5-membered heteroaromaticring containing four nitrogen atoms is tetrazolyl. Suitable 6-memberedheteroaromatic rings containing three nitrogen atoms include1,2,4-triazine and 1,3,5-triazine. When a heteroaromatic ring comprisesa hydroxy group as a substituent, and keto-enol tautomerism is possible,both tautomers are included within the scope of the invention. Thus, forexample, a 3-hydroxy-1,2,4-triazole ring will be considered equivalentto the 3-keto tautomer.

[0030] The expression “heteroaryl(C₁₋₆)alkyl” as used herein includesfurylmethyl, furylethyl, thienylmethyl, thienylethyl, pyrazolylmethyl,oxazolylmethyl, oxazolylethyl, isoxazolylmethyl, thiazolylmethyl,thiazolylethyl, imidazolylmethyl, imidazolylethyl, benzimidazolylmethyl,oxadiazolylmethyl, oxadiazolylethyl, thiadiazolylmethyl,thiadiazolylethyl, triazolylmethyl, triazolylethyl, tetrazolylmethyl,tetrazolylethyl, pyridinylmethyl, pyridinylethyl, pyridazinylmethyl,pyrimidinylmethyl, pyrazinylmethyl, quinolinylmethyl,isoquinolinylmethyl and quinoxalinylmethyl.

[0031] The term “halogen” as used herein includes fluorine, chlorine,bromine and iodine, of which fluorine and chlorine are preferred.

[0032] As used herein the term “C₁₋₆alkoxy” includes methoxy and ethoxygroups, and straight-chained, branched and cyclic propoxy, butoxy,pentoxy and hexoxy groups, including cyclopropylmethoxy. Derivedexpressions such as “C₂₋₆alkenyloxy”, “C₂₋₆alkynyloxy”, “C₁₋₄alkoxy”,“C₂₋₄alkenyloxy” and “C₂₋₄alkyloxy” should be construed in an analogousmanner.

[0033] For use in medicine, the salts of the compounds of formula I willbe pharmaceutically acceptable salts. Other salts may, however, beuseful in the preparation of the compounds according to the invention orof their pharmaceutically acceptable salts. Suitable pharmaceuticallyacceptable salts of the compounds of this invention include acidaddition salts which may, for example, be formed by mixing a solution ofthe compound according to the invention with a solution of apharmaceutically acceptable acid such as hydrochloric acid, sulphuricacid, methanesulphonic acid, fumaric acid, maleic acid, succinic acid,acetic acid, benzoic acid, oxalic acid, citric acid, tartaric acid,carbonic acid or phosphoric acid. Furthermore, where the compounds ofthe invention carry an acidic moiety, suitable pharmaceuticallyacceptable salts thereof may include alkali metal salts, e.g. sodium orpotassium salts; alkaline earth metal salts, e.g. calcium or magnesiumsalts; and salts formed with suitable organic ligands, e.g. quaternaryammonium salts.

[0034] Where the compounds according to the invention have at least oneasymmetric centre, they may accordingly exist as enantiomers. Where thecompounds according to the invention possess two or more asymmetriccentres, they may additionally exist as diastereoisomers. It is to beunderstood that all such isomers and mixtures thereof in any proportionare encompassed within the scope of the present invention.

[0035] Preferably L is an oxygen atom. L may also be S or NR^(n) inwhich R^(n) is preferably hydrogen or methyl. R^(n) may be hydrogen.

[0036] X is generally: pyridyl, pyrazinyl, pyridazinyl or pyrimidinyloptionally substituted by a halogen atom or a group R³, OR³, NR⁴R⁵ or afive membered heteroaromatic ring containing 1, 2 or 3 nitrogen atoms,and X is optionally fused to a benzene ring; a 5-membered heteroaromaticring containing 2 or 3 heteroatoms chosen from oxygen, sulphur andnitrogen, at most one of the heteroatoms being oxygen or sulphur, whichis unsubstituted or substituted by one, two or three groupsindependently chosen from halogen and R³, or which is substituted by apyridyl, phenyl or benzyl ring which ring is optionally independentlysubstituted by one, two or three halogen atoms or C₁₋₆alkyl or CF₃groups; or phenyl optionally substituted by one, two or threeindependently chosen halogen atoms.

[0037] Alternatively, X may represent a 5-membered heteroaromatic ringcontaining 2 or 3 heteroatoms chosen from oxygen, sulphur and nitrogen,at most one of the heteroatoms being oxygen or sulphur, which is fusedto a benzene or pyridine ring, or which is substituted by NH₂ or by OHwhich may exist as the keto tautomer; or X may represent a tetrazolering bearing a C₁₋₄alkyl substituent.

[0038] When X is a substituted 6-membered heteroaromatic ring: R^(x) ispreferably halogen, R³, OR³, NR⁴R⁵ or a five-membered heteroaromaticring containing 1, 2 or 3 nitrogen atoms and more preferably methyl,CF₃, methoxy, bromine, chlorine, isopropoxy, dimethylamino or afive-membered heterocyclic ring containing 1, 2 or 3 nitrogen atoms; andR^(y) and R^(z) are preferably absent.

[0039] When X is a substituted 5-membered heteroaromatic ring: R^(x) ispreferably halogen, R³ or a pyridyl, phenyl or benzyl ring, which ringis optionally independently substituted by one, two or three halogenatoms or C₁₋₆alkyl or CF₃ groups, and more preferably R^(x) is methyl,CF₃, chlorine or a phenyl, pyridyl or benzyl ring which ring isunsubstituted or substituted by chlorine or CF₃; and R^(y) and R^(z) arepreferably halogen or R³ or are absent, and more preferably are methyl,CF₃ or chlorine, or are absent.

[0040] X is preferably a substituted triazolyl, such as2-methyl-1,2,4-triazol-5yl.

[0041] X is preferably pyridyl or substituted pyridyl, such aspyrid-2-yl and 3-methoxypyrid-2-yl.

[0042] Apt values for Y include CH₂, CH(CH₃), CH₂CH₂ and CH₂CH₂CH₂optionally substituted by an oxo group, and CH₂CH₂O and CH₂CH₂CH₂O. Forexample, Y can be CH₂, CH₂CH₂, CH₂CH₂CH₂, CH₂CH₂O or CH₂CH₂CH₂O.Preferably Y is CH₂ or CH₂CH₂ and most preferably CH₂.

[0043] From the foregoing it will be understood that particularlysuitable groups L-Y—X are OCH₂X groups where X is pyridyl or substitutedpyridyl.

[0044] Suitable values for Z include optionally substituted phenyl,pyrimidinyl, triazinyl, pyrrolyl, oxazolyl, isoxazolyl, thiazolyl,isothiazolyl, oxadiazolyl and thiadiazolyl groups.

[0045] Z is very aptly an optionally substituted 5-memberedheteroaromatic ring containing one oxygen and one or two nitrogen ringatoms.

[0046] Favoured values for Z include optionally substituted phenyls andisoxazoles.

[0047] Typical substituents on Z include C₁₋₆alkyl, C₃₋₆cycloalkyl,hydroxyC₁₋₆alkyl, pyridyl, thienyl or amino, particularly methyl, ethyl,isopropyl, cyclopropyl, thienyl or pyridyl.

[0048] Z may be unsubstituted.

[0049] Z may very aptly be substituted by methyl.

[0050] Particular values of Z include 5-methylisoxazol-3-yl and phenyl.Z may also be isoxazol-3-yl.

[0051] A particular sub-class of compounds according to the invention isrepresented by the compounds of formula IIA, and pharmaceuticallyacceptable salts thereof.

[0052] wherein

[0053] X′ represents, pyrazolyl, isoxazolyl, thiazolyl, imidazolyl,thiadiazolyl, benzimidazolyl, oxadiazolyl, triazolyl, tetrazolyl,pyridinyl, pyridazinyl, oxazolopyridinyl, pyrimidinyl, pyrazinyl,quinolinyl, isoquinolinyl or quinoxalinyl, any of which groups may beoptionally substituted by one or more of C₁₋₆ alkyl, amino, pyridyl,CF₃, aryl(C₁₋₆)alkyl, pyridyl(C₁₋₆)alkyl, halogen, cyano,cyano(C₁₋₆)alkyl, hydroxymethyl, C₁₋₆alkoxy, C₃₋₆cycloalkyl(C₁₋₆)alkoxy,hydroxy or the keto tautomer thereof, di(C₁₋₆)alkylamino(C₁₋₆)alkyl,amino(C₁₋₆)alkyl, N-(C₁₋₆)alkylpiperidinyl, pyrrolidinyl(C₁₋₆)alkyl,piperazinyl(C₁₋₆)alkyl and morpholinyl(C₁₋₆)alkyl; and

[0054] Z′ represents a phenyl ring or a 5-membered heteroaromatic ringcontaining one oxygen and one or two nitrogen ring atoms each of whichrings is optionally substituted with C₁₋₆alkyl or C₁₋₆alkoxy.

[0055] Particular compounds within the scope of the present inventioninclude:

[0056] 6-(2-pyridy)methyloxy-3-phenylimidazo[5,1-a]phthalazine;

[0057]6-(3-methoxypyridin-2-yl)methyloxy-3-(5-methylisoxazol-3-yl)imidazo[5,1-a]phthalazine;

[0058] and pharmaceutically acceptable salts thereof The compounds ofthe present invention have a good binding affinity (Ki) for the α5subunit of the GABA_(A) receptor. In a preferred embodiment thecompounds of the invention are binding selective for the α5 subunitrelative to the α1, α2 and α3 subunits. In another preferred embodimentthe compounds are functionally selective for the α5 subunit as partialor full inverse agonists whilst substantially being antagonists at theα1, α2 and α3 subunits.

[0059] The invention also provides pharmaceutical compositionscomprising one or more compounds of this invention and apharmaceutically acceptable carrier. Preferably these compositions arein unit dosage forms such as tablets, pills, capsules, powders,granules, sterile parenteral solutions or suspensions, metered aerosolor liquid sprays, drops, ampoules, transdermal patches, auto-injectordevices or suppositories; for oral, parenteral, intranasal, sublingualor rectal administration, or for administration by inhalation orinsufflation. For preparing solid compositions such as tablets, theprincipal active ingredient is mixed with a pharmaceutical carrier, e.g.conventional tableting ingredients such as corn starch, lactose,sucrose, sorbitol, talc, stearic acid, magnesium stearate, dicalciumphosphate or gums or surfactants such as sorbitan monooleate,polyethylene glycol, and other pharmaceutical diluents, e.g. water, toform a solid preformulation composition containing a homogeneous mixtureof a compound of the present invention, or a pharmaceutically acceptablesalt thereof. When referring to these preformulation compositions ashomogeneous, it is meant that the active ingredient is dispersed evenlythroughout the composition so that the composition may be readilysubdivided into equally effective unit dosage forms such as tablets,pills and capsules. This solid preformulation composition is thensubdivided into unit dosage forms of the type described above containingfrom 0.1 to about 500 mg of the active ingredient of the presentinvention. Typical unit dosage forms contain from 1 to 100 mg, forexample 1, 2, 5, 10, 25, 50 or 100 mg, of the active ingredient. Thetablets or pills of the novel composition can be coated or otherwisecompounded to provide a dosage form affording the advantage of prolongedaction. For example, the tablet or pill can comprise an inner dosage andan outer dosage component, the latter being in the form of an envelopeover the former. The two components can be separated by an enteric layerwhich serves to resist disintegration in the stomach and permits theinner component to pass intact into the duodenum or to be delayed inrelease. A variety of materials can be used for such enteric layers orcoatings, such materials including a number of polymeric acids andmixtures of polymeric acids with such materials as shellac, cetylalcohol and cellulose acetate.

[0060] The present invention also provides a compound of the inventionfor use in a method of treatment of the human body. Preferably thetreatment is for a condition associated with GABA_(A) receptorscomprising the α5 subunit and/or for the enhancement of cognition.Preferably the condition is a neurological deficit with an associatedcognitive disorder such as a dementing illness such as Alzheimer'sdisease. Other conditions to be treated include cognition deficits dueto traumatic injury, stroke, Parkinson's disease, Downs syndrome, agerelated memory deficits, attention deficit disorder and the like.

[0061] The present invention further provides the use of a compound ofthe present invention in the manufacture of a medicament for theenhancement of cognition, preferably in a human suffering from adementing illness such as Alzheimer's disease.

[0062] Also disclosed is a method of treatment of a subject sufferingfrom a cognition deficit, such as that resulting from a dementingillness such as Alzheimer's disease, which comprises administering tothat subject an effective amount of a compound according to the presentinvention.

[0063] Thus, the compounds of the present invention can be used in avariety of disorders of the central nervous system. Such disordersinclude delirium, dementia and amnestic and other cognitive disorders.Examples of delirium are delirium due to substance intoxication orsubstance withdrawal, delirium due to multiple etiologies and deliriumNOS (not otherwise specified). Examples of dementia are: dementia of theAlzheimer's type with early onset which can be uncomplicated or withdelirium, delusions or depressed mood; dementia of the Alzheimer's type,with late onset, which can be uncomplicated or with delirium, delusionsor depressed mood; vascular dementia which can be uncomplicated or withdelirium, delusions or depressed mood; dementia due to HIV disease;dementia due to head trauma; dementia due to Parkinson's disease;dementia due to Huntington's disease; dementia due to Pick's disease;dementia due to Creutzfeld-Jakob disease; dementia which issubstance-induced persisting or due to multiple etiologies; and dementiaNOS. Examples of amnestic disorders are amnestic disorder due to aparticular medical condition or which is substance-induced persisting orwhich is amnestic disorder NOS.

[0064] The liquid forms in which the novel compositions of the presentinvention may be incorporated for administration orally or by injectioninclude aqueous solutions, suitably flavoured syrups, aqueous or oilsuspensions, and flavoured emulsions with edible oils such as cottonseedoil, sesame oil, coconut oil or peanut oil, as well as elixirs andsimilar pharmaceutical vehicles. Suitable dispersing or suspendingagents for aqueous suspensions include synthetic and natural gums suchas tragacanth, acacia, alginate, dextran, sodium carboxymethylcellulose,methylcellulose, polyvinyl-pyrrolidone or gelatin.

[0065] For the enhancement of cognition, a suitable dosage level isabout 0.01 to 250 mg/kg per day, preferably about 0.01 to 100 mg/kg perday, and especially about 0.01 to 5 mg/kg of body weight per day. Thecompounds may be administered on a regimen of 1 to 4 times per day. Insome cases, however, dosage outside these limits may be used.

[0066] There is also provided a process for producing a compound offormula I which comprises reacting a compound of formula II with acompound of formula III:

[0067] wherein L, X, Y and Z are as defined above, generally in thepresence of a base such as sodium hydride or lithiumbis(trimethylsilyl)amide, and a solvent such as a mixture oftetrahydrofuran and N,N-dimethylformamide at a temperature of between−78° C. and room temperature.

[0068] The compounds of formula II can be made by reacting a compound offormula IV with a chlorinating agent such as POCl₃:

[0069] wherein Z is as defined above, generally at elevated temperature,such as from 85-120° C., for about 0.5 to 2.5 hours, optionally followedby heating at reflux with a catalyst such as Et₃N.HCl in a solvent suchas xylene.

[0070] The compounds of formula IV can be produced by reacting acompound of formula V with 4-aminomethyl-2H-phthalazin-1-onehydrochloride (Geterotsikl. Soedin., 1972, (9), 15):

[0071] wherein Z is as defined above generally in the presence of a basesuch as triethylamine and a solvent such as dimethylformamide.

[0072] Compounds of formulae III and V are commercially available or canbe made from commercially available compounds by known methods.

[0073] It will be understood that any compound of formula I initiallyobtained from any of the above processes may, where appropriate,subsequently be elaborated into a further compound of formula I bytechniques known from the art.

[0074] It will also be appreciated that where more than one isomer canbe obtained from a reaction then the resulting mixture of isomers can beseparated by conventional means.

[0075] Where the above-described process for the preparation of thecompounds according to the invention gives rise to mixtures ofstereoisomers, these isomers may be separated by conventional techniquessuch as preparative chromatography. The novel compounds may be preparedin racemic form, or individual enantiomers may be prepared either byenantiospecific synthesis or by resolution. The novel compounds may, forexample, be resolved into their component enantiomers by standardtechniques such as preparative HPLC, or the formation of diastereomericpairs by salt formation with an optically active acid, such as(−)-di-p-toluoyl-d-tartaric acid and/or (+)-di-p-toluoyl-l-tartaricacid, followed by fractional crystallization and regeneration of thefree base. The novel compounds may also be resolved by formation ofdiastereomeric esters or amides, followed by chromatographic separationand removal of the chiral auxiliary.

[0076] During any of the above synthetic sequences it may be necessaryand/or desirable to protect sensitive or reactive groups on any of themolecules concerned. This may be achieved by means of conventionalprotecting groups, such as those described in Protective Groups inOrganic Chemistry, ed. J. F. W. McOmie, Plenum Press, 1973; and T. W.Greene & P. G. M. Wuts, Protective Groups in Organic Synthesis, JohnWiley & Sons, 1991. The protecting groups may be removed at a convenientsubsequent stage using methods known from the art.

[0077] The compounds in accordance with this invention potently inhibitthe binding of [3H]-flumazenil to the benzodiazepine binding site ofhuman GABA_(A) receptors containing the α5 subunit stably expressed inLtk-cells.

[0078] Reagents

[0079] Phosphate buffered saline (PBS).

[0080] Assay buffer: 10 mM KH₂PO₄, 100 mM KCl, pH 7.4 at roomtemperature.

[0081] [3H]-Flumazenil (18 nM for α1β3γ2 cells; 18 nM for α2β3γ2 cells;10 nM for α3β3γ2 cells; 10 nM for α5β3γ2 cells) in assay buffer.

[0082] Flunitrazepam 100 μM in assay buffer.

[0083] Cells resuspended in assay buffer (1 tray to 10 ml).

[0084] Harvesting Cells

[0085] Supernatant is removed from cells. PBS (approximately 20 ml) isadded. The cells are scraped and placed in a 50 ml centrifuge tube. Theprocedure is repeated with a further 10 ml of PBS to ensure that most ofthe cells are removed. The cells are pelleted by centrifuging for 20 minat 3000 rpm in a benchtop centrifuge, and then frozen if desired. Thepellets are resuspended in 10 ml of buffer per tray (25 cm×25 cm) ofcells.

[0086] Assay

[0087] Can be carried out in deep 96-well plates or in tubes. Each tubecontains:

[0088] 300 μl of assay buffer.

[0089] 50 μl of [3H]-flumazenil (final concentration for α1β3γ2: 1.8 nM;for α2β3γ2: 1.8 nM; for α3β3γ2: 1.0 nM; for α5β3γ2: 1.0 nM).

[0090] 50 μl of buffer or solvent carrier (e.g. 10% DMSO) if compoundsare dissolved in 10% DMSO (total); test compound or flunitrazepam (todetermine non-specific binding), 10 μM final concentration.

[0091] 100 μl of cells.

[0092] Assays are incubated for 1 hour at 40° C., then filtered usingeither a Tomtec or Brandel cell harvester onto GF/B filters followed by3×3 ml washes with ice cold assay buffer. Filters are dried and countedby liquid scintillation counting. Expected values for total binding are3000-4000 dpm for total counts and less than 200 dpm for non-specificbinding if using liquid scintillation counting, or 1500-2000 dpm fortotal counts and less than 200 dpm for non-specific binding if countingwith meltilex solid scintillant. Binding parameters are determined bynon-linear least squares regression analysis, from which the inhibitionconstant Ki can be calculated for each test compound.

[0093] The compounds of the accompanying Examples were tested in theabove assay, and all were found to possess a Ki value for displacementof [3H]Ro 15-1788 from the α5 subunit of the human GABA_(A) receptor of100 nM or less, most were at 50 nM or less, many were at 10 nM or lessand some were at 1 nM or less.

[0094] The compounds of the present invention have been shown to enhancecognition in the rat water maze test (Morris, Learning and Motivation,1981, 12, 239ff). Further details of methodology for demonstrating thatthe present compounds enhance cognition can be found in WO-A-9625948.

[0095] The following Examples illustrate the present invention:

EXAMPLE 1 6-(2-Pyridyl)methyloxy-3-phenylimidazo[5,1-a]phthalazine a)N-(4-Oxo-3,4-dihydrophthalazin-1-ylmethyl)benzamide

[0096] Triethylamine (5.9 ml, 42 mmol) was added dropwise to a stirredsuspension of 4-aminomethyl-2H-phthalazin-1-one hydrochloride(Geterotsikl. Soedin, 1972, (9), 15) (3.00 g, 14.2 mmol) in DMF (150 ml)at room temperature under nitrogen. The mixture was stirred for 1.5 hbefore the addition of benzoyl chloride (2.5 ml, 22 mmol) and thenstirred at room temperature overnight. The reaction mixture was filteredand water added to the filtrate to form a solid. The mixture was cooledand the solid collected by filtration, washed successively with diethylether and isohexane, and dried in vacuo to give the title-compound (3.18g, 80%), ¹H NMR (360 MHZ, d⁶-DMSO) δ 4.81 (2H, d, J=5.6 Hz, CH₂),7.43-7.53 (3H, m, 3 of Ar—H), 7.83-7.96 (4H, m, 4 of Ar—H), 8.08 (1H, d,J=7.9 Hz, Ar—H), 8.28 (1H, dd, J=7.8 and 1.2 Hz, Ar—H), 8.98 (1H, br s,NH), 12.62 (1H, s NH).

b) 6-Chloro-3-phenylimidazo[5,1-a]phthalazine

[0097] A suspension of the preceding product (2.48 g, 8.88 mmol) inphosphorus oxychloride (118 ml) was heated at reflux under nitrogen for2.5 h. The solvent was evaporated in vacuo and the residue azeotropedwith toluene, diluted with dichloromethane and water and the mixturebasified with solid sodium carbonate. The layers were separated and theaqueous layer was extracted with dichloromethane (×2). The combinedorganic layers were dried (MgSO₄) and evaporated in vacuo and theresidue purified by chromatography on silica gel, eluting with 1→3%methanol/dichloromethane (gradient elution), to give the title-compound(1.00 g, 40%), ¹H NMR (360 MHz, CDCl₃) δ 7.41-7.59 (4H, m, 4 of Ar—H),7.79 (1H, t, J=7.6 Hz, Ar—H), 7.88 (1H, s, Ar—H), 8.02 (1H, d, J=7.7 Hz,Ar—H), 8.12 (1H, m, Ar—H), 8.39-8.41 (2H, m, 2 of Ar—H).

c) 6-(2-Pyridyl)methyloxy-3-phenylimidazo[5,1-a]phthalazine

[0098] Sodium hydride (30 mg of a 60% dispersion, 0.75 mmol) was addedto a stirred solution of the preceding product (116 mg, 0.41 mmol) and2-pyridylcarbinol (0.080 ml, 0.83 mmol) in DMF (5 ml) at roomtemperature under nitrogen. After 1.5 h, water was added and the mixturewas evaporated in vacuo. The residue was partitioned betweendichloromethane and water and the aqueous layer was separated andfurther extracted with dichloromethane (×2). The combined organicextracts were dried (Na₂SO₄) and evaporated in vacuo and the residuepurified by chromatography on silica gel, eluting with 20% ethylacetate/dichloromethane, followed by recrystallisation (ethylacetate/hexane) to give the title-product (70 mg, 48%), ¹H NMR (250 MHz,CDCl₃) δ 5.70 (2H, s, CH₂), 7.30-7.59 (6H, m, 6 of Ar—H), 7.72-7.76 (2H,m, 2 of Ar—H), 7.82 (1H, s, Ar—H), 7.99 (1H, d, J=7.9 Hz, Ar—H), 8.18(1H, d, J=8.0 Hz, Ar—H), 8.29-8.34 (2H, m, 2 of Ar—H), 8.70 (1H, br d,Ar—H); MS (ES⁺) m/e 353 [MH]⁺.

EXAMPLE 26-(3-Methoxypyridin-2-yl)methyloxy-3-(5-methylisoxazol-3-yl)imidazo[5,1-a]phthalazinea) 5-Methylisoxazole-3-carboxylic acid(4-oxo-3,4-dihydroiohthalazin-1-ylmethyl)amide

[0099] The title compound was prepared from4-aminomethyl-2H-phthalazine-1-one hydrochloride (Geterotsikl. Soedin,1972, (9), 15) and 5-methylisoxazole-3-carbonyl chloride using theprocedure described in Example 1, Part a). ¹H NMR (360 MHz, d⁶-DMSO) δ2.45 (3H, d, J=0.6 Hz, CH₃), 4.77 (2H, d, J=5.8 Hz, CH₂), 6.57 (1H, d,J=0.7 Hz, Ar—H), 7.86 (1H, t, J=8.2 Hz, Ar—H), 7.94.(1H, t, J=8.1 Hz,Ar—H), 8.06 (1H, d, J=7.6 Hz, Ar—H), 8.28 (1H, d, J=8.0 Hz, Ar—H), 9.12(1H, t, J=5.7 Hz, NH) and 12.61 (1H, S, NH).

b) 6-Chloro-3-(5-methylisoxazol-3-yl)-imidazo[5,1-a]phthalazine

[0100] The amide (3.2 g, 0.011 mol) was dissolved in phosphorusoxychloride (120 ml) and heated at 90° C. for 0.5 h under nitrogen. Thesolvent was removed in vacuo and the residue azeotroped with toluene(2×). The resulting brown oil was dissolved in xylene (120 ml) andheated at reflux for 1 h. The solvent was removed in vacuo and theresidue purified by chromatography on silica gel, eluting with 2%methanol/dichloromethane. Further chromotographic purification elutingwith 20% ethyl acetate/dichloromethane gave the title-compound as ayellow solid (0.65 g, 20%), ¹H NMR (360 MHz, CDCl₃) δ 2.55 (3H, d, J=0.8Hz, CH₃), 6.87 (1H, d, J=0.7 Hz, Ar—H), 7.64 (1H, t, J=7.4 H₂, Ar—H),7.85 (1H, t, J=7.4 Hz, Ar—H), 7.95 (1H, s, Ar—H), 8.06 (1H, d, J=8.1 Hz,Ar—H), 8.18 (1H, d, J=8.1 Hz, Ar—H); MS (ES⁺) m/e 284 [MH]⁺.

c) (3-Methoxypyridin-2-yl)methanol

[0101] Dimethylsulphoxide (15 ml) was added to ground potassiumhydroxide (10.2 g, 0.186 mol) and stirred for 0.25 h at room temperatureunder nitrogen. 3-Hydroxy-(2 hydroxymethyl)pyridine hydrochloride (5.0g, 0.031 mol) in dimethylsulphoxide (20 ml) was added via syringe. Afterstirring for 1 h, methyl iodide (1.92 ml, 0.031 mol) was added and thereaction stirred at room temperature overnight. Water (100 ml) was addedand the mixture acidified to pH1 with 5N hydrochloric acid. The aqueouslayer was washed with dichloromethane (×3), basified to pH14 with sodiumhydroxide and extracted with dichloromethane (×4). The combined organicextracts were dried (MgSO₄), the solvent removed in vacuo and theresidue purified by chromatography eluting withdichloromethane/methanol/ammonia (90:5:0.5), to give the title-compound,¹H NMR (360 MHz, CDCl₃) δ 3.86 (3H, d, J=1.8 Hz, CH₃), 4.29 (¹H, t,J=4.5 Hz, OH), 4.74 (2H, d, J=4.1 Hz, CH₂), 7.12-7.22 (2H, m, 2 of Ar—H)and 8.15 (1H, d, J=4.7 Hz, Ar—H).

d)6-(3-Methoxypyridin-2-yl)methyloxy-3-(5-methylisoxazol-3-yl)imidazo[5,1-a]phthalazine

[0102] To the preceding alcohol (52 mg, 0.38 mmol) dissolved inN,N-dimethylformamide (1 ml) and tetrahydrofuran (2.5 ml) was added theproduct of part b above (100 mg, 0.35 mmol) and the mixture stirred at−78° C. under nitrogen for 0.5 h. Lithium bis(trimethylsilyl)amide (0.42ml of 1.0 M solution in hexanes, 0.42 mmol) was added and the reactionallowed to warm up gradually to room temperature overnight. The reactionmixture was then heated at 50° C. for 5 h. The solvent was removed invacuo and the residue azeotroped with xylene (×2) and purified bychromatography on silica gel, eluting with 0→5%methanol/dichloromethane, followed by recrystallisation(dichloromethane/hexane) to give the title-compound (41 mg, 32%) as apale yellow solid, ¹H NMR (360 MHz, CDCl₃) δ 2.52 (3H, s, CH₃), 3.89(3H, s, CH₃), 5.75 (2H, s, CH₂), 6.87 (1H, s, Ar—H), 7.26-7.34 (2H, m, 2of Ar—H), 7.48 (1H, t, J=7.3 Hz, Ar—H), 7.74 (1H, t, J=7.3 Hz, Ar—H),7.86 (1H, d, J=0.6 Hz, Ar—H), 8.00 (1H, d, J=8.1 Hz, Ar—H), 8.13 (1H, d,J=8.1 Hz, Ar—H), 8.25 (1H, d, J=4.4 Hz, Ar—H); MS (ES⁺) m/e 388 [MH]⁺.

EXAMPLE 36-(2-Methyl-1,2,4-triazol-5-yl)methyloxy-3-(isoxazol-3-yl)imidazo[5,1-a]phthalazinea) Isoxazol-3-carboxylic acid(4-oxo-3,4-dihydrophthalazin-1-ylmethyl)amide

[0103] The title compound was prepared from4-aminomethyl-2H-phthalazine-1-one hydrochloride (Geterotsikl, Soedin,1972, (9), 15) and isoxazol-3-carbonyl chloride using the proceduredescribed in Example 1, Part a. ¹H NMR (360 MHz, d⁶-DMSO) δ 4.80 (d, J=6Hz, 2H), 6.93 (s, 1H), 7.87 (t, J=7 Hz, 1H), 7.95 (t, J=7 Hz, 1H), 8.1(d, J=7 Hz, 1H), 8.28 (dd, J=2 Hz, 7 Hz, 1H), 9.08 (s, 1H), 9.09 (t, J=2Hz, 1H), 12.62 (s, 1H).

b) 6-Chloro-3-(isoxazol-3-yl)-imidazo[5,1-a]phthalazine

[0104] The title compound was prepared from isoxazol-3-carboxylic acid(4-oxo-3,4-dihydrophthalazin-1-ylmethyl)amide (Part a) and phosphorusoxychloride according the procedure described in Example 1, Part b. ¹HNMR (360 MHz, CDCl₃) δ 7.25 (s, 1H), 7.66 (t, J=8 Hz, 1H), 7.86 (t, J=8Hz, 1H), 7.97 (brs, 1H), 8.09 (d, J=8 Hz, 1H), 8.21 (d, J=8 Hz, 1H),8.57 (d, J=2 Hz, 1H).

c)6-(2-Methyl-1,2,4-triazol-5-yl)methyloxy-3-(isoxazol-3-yl)imidazo[5,1-a]phthalazine

[0105] The title compound was prepared from6-chloro-3-(isoxazol-3-yl)-imidazo[5,1-a]phthalazine (Part b) and(2-methyl-1,2,4-triazol-5-yl)methanol as described in Example 1, Part c.¹H NMR (360 MHz, CDCl₃) δ 3.96 (s, 3H), 5.7 (s, 2H), 7.40 (s, 1H), 7.52(t, J=8 Hz, 1H), 7.76 (t, J=8 Hz, 1H), 7.89 (s, 1H), 8.0 (d, J=8 Hz,1H), 8.17 (d, J=8 Hz, 1H), 8.56 (s, 1H). MS (ES⁺) m/e 348 [MH]⁺.

1. A compound of formula I, or a pharmaceutically acceptable salt thereof.

wherein L is O, S or NR^(n) where R^(n) is H, C₁₋₆alkyl or C₃₋₆cycloalkyl; X is a 5-membered heteroaromatic ring containing 1, 2, 3 or 4 heteroatoms independently chosen from oxygen, nitrogen and sulphur, at most one of the heteroatoms being oxygen or sulphur, or a 6-membered heteroaromatic ring containing 1, 2 or 3 nitrogen atoms, the 5- or 6-membered heteroaromatic ring being optionally fused to a benzene or pyridine ring and the heteroaromatic ring being optionally substituted by R^(x) and/or R^(y) and/or R^(z), where R^(x) is halogen, R³, OR³, OCOR³, NR⁴R⁵, NR⁴COR⁵, OH, tri(C₁₋₆alkyl)silylC₁₋₆alkoxyC₁₋₄alkyl, CN or R⁶, R^(y) is halogen, R³, OR³, OCOR³, NR⁴R⁵, NR⁴COR⁵, NR⁴R⁵(C₁₋₆)alkyl or CN and R^(z) is R³, OR³ or OCOR³, providing that when X is a pyridine derivative, the pyridine ring is optionally in the form of the N-oxide, and providing that when X is a tetrazole derivative it is protected by a C₁₋₄alkyl group; Y is optionally branched C₁₋₄alkylene optionally substituted by an oxo group or Y is a group (CH₂)_(j)O wherein the oxygen atom is nearest the group X and j is 2, 3 or 4; Z represents a phenyl ring, a 5-membered heteroaromatic ring containing 1, 2 or 3 heteroatoms independently selected from oxygen, nitrogen and sulphur, at most one of the heteroatoms being oxygen or sulphur, or a 6-membered heteroaromatic ring containing 1, 2 or 3 nitrogen atoms with the exception of pyrazine, any of which rings may be optionally substituted with one or more substituents selected from halogen, R³, OR³, OCOR³, NR⁴R⁵, NR⁴R⁵(C₁₋₆)alkyl, NR⁴R⁵CO, NR⁴R⁵CO(C₁₋₆)alkyl, CN, cyano(C₁₋₆)alkyl or R⁶; R³ represents C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₁₋₆cycloalkyl, C₃₋₆cycloalkyl(C₁₋₆)allyl, cyano(C₁₋₆)alkyl, hydroxyC₁₋₆alkyl and R³ is optionally mono, di- or tri-fluorinated; R⁴ and R⁵ are each independently hydrogen, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkyl, C₁₋₆cycloalkyl or CF₃ or R⁴ and R^(5,) together with a nitrogen atom to which they are commonly attached, form a 4-7 membered heteroaliphatic ring containing the said nitrogen atom and optionally one other heteroatom selected from O, N and S; and R⁶ is C₆₋₁₀aryl, C₆₋₁₀aryl (C₁₋₆)alkyl, heteroaryl or heteroaryl(C₁₋₆)alkyl, where heteroaryl is a five-membered aromatic ring containing one, two, three or four nitrogen atoms or one oxygen atom or sulphur atom and optionally one or two nitrogen atoms or a six-membered aromatic ring containing one, two or three nitrogen atoms, and R⁶ is optionally substituted by one, two or three substituents independently chosen from halogen atoms and C₁₋₄alkyl, C₂₋₄alkenyl, C₂₋₄alkynyl, C₁₋₄alkoxy, C₂₋₄alkenyloxy and C₂₋₄ alkynyloxy groups each of which groups is unsubstituted or substituted by one, two or three halogen atoms.
 2. A compound as claimed in claim 1 represented by a compound of formual IIA, or a pharmaceutically acceptable salt thereof.

wherein X′ represents, pyrazolyl, isoxazolyl, thiazolyl, imidazolyl, thiadiazolyl, benzimidazolyl, oxadiazolyl, triazolyl, tetrazolyl, pyridinyl, pyridazinyl, oxazolopyridinyl, pyrimidinyl, pyrazinyl, quinolinyl, isoquinolinyl or quinoxalinyl, any of which groups may be optionally substituted by one or more of C₁₋₆ alkyl, amino, pyridyl, CF₃, aryl(C₁₋₆)alkyl, pyridyl(C₁₋₆)alkyl, halogen, cyano, cyano(C₁₋₆)alkyl, hydroxymethyl, C₁₋₆alkoxy, C₃₋₆cycloalkyl(C₁₋₆)alkoxy, hydroxy or the keto tautomer thereof, di(C₁₋₆)alkylamino(C₁₋₆)alkyl, amino(C₁₋₆)alkyl, N—(C₁₋₆)alkylpiperidinyl, pyrrolidinyl(C₁₋₆)alkyl, piperazinyl(C₁₋₆)alkyl and morpholinyl(C₁₋₆)alkyl; and Z′ represents a phenyl ring or a 5-membered heteroaromatic ring containing one oxygen and one or two nitrogen ring atoms each of which rings is optionally substituted with C₁₋₆alkyl or C₁₋₆alkoxy.
 3. A compound selected from: 6-(2-pyridyl)methyloxy-3-phenylimidazo[5,1-a]phthalazine; 6-(3-methoxypyridin-2-yl)methyloxy-3-(5-methylisoxazol-3-yl)imidazo[5,1-a]phthalazine; 6-(2-Methyl-1,2,4-triazol-5-yl)methyloxy-3-(isoxazol-3-yl)imidazo[5,1-a]phthalazine; and pharmaceutically acceptable salts thereof.
 4. A pharmaceutical composition comprising a compound of formula I as defined in claim 1, 2 or 3 or a pharmaceutically acceptable salt thereof in combination with a pharmaceutically acceptable carrier.
 5. The use of a compound as claimed in any one of claims 1 to 3 for the manufacture of a medicament for the enhancement of cognition.
 6. A process for the preparation of a compound as claimed in claim 1 which comprises: (A) reacting a compound of formula II with a compound of formula III:

wherein L, X, Y and Z are as defined in claim 1; and (B) subsequently, if desired, converting a compound of formula I initially obtained into a further compound of formula I by techniques known from the art.
 7. A method of treatment of a subject suffering from a cognition deficit which comprises administering to that subject an effective amount of a compound of formula I as defined in claim 1, or a pharmaceutically acceptable salt thereof. 